Project Summary/Abstract (30 lines) Ascending thoracic aortic aneurysms and dissections (ATAAD) are extremely lethal surgical diseases. ATAAD occurs either in association with genetic conditions or spontanousely as sporadic ATAAD, which accounts for more than 80% of ATAAD. Unfortunately, no clinically proven medication is available to prevent sporadic ATAAD progression. There is a critical need to develop effective pharmacological strategies to treat these dis- eases. This project focuses on sporadic ATAAD. One of the significant features of sporadic ATAAD is progres- sive smooth muscle cell (SMC) depletion, for which the underlying mechanisms are poorly understood. Ne- crotic cell death, a type of cell death characterized by cell swelling and membrane rupture, causes the re- lease of ?toxic? molecules from dying cells, leading to further damage of surrounding cells and extracellular ma- trix and a profound pro-inflammatory reaction. Necrotic cell death is regulated by a programmed process called necroptosis. Mitochondrial damage, often observed in aged or stressed cells, is a potent trigger for necrotic cell death. The overall objective of this application is to determine the role of mitochondrial damage and SMC necroptosis in the development of TAAD; and the extent to which this mechanism represents a therapeutic tar- get against ATAAD. Our central hypotheses are: (1) mitochondrial damage and subsequent release of mtDNA trigger the cytosolic DNA sensor cGAS-STING-TBK1 signaling pathway, which activates the RIP3-MLKL path- way and induces SMC necroptosis, contributing to aortic damage and ATAAD formation; and (2) reducing mi- tochondrial damage or inhibiting SMC necroptosis will prevent aortic destruction and ATAAD progression. We will test these hypotheses via 3 aims. Aim 1. We will test the hypothesis that mitochondrial damage and sub- sequent release of mtDNA activates the cytosolic DNA sensor cGAS-STING-TBK1 signaling pathway, which in turn activates the RIP3-MLKL necroptosis pathway, leading to SMC necrotic death. We will perform serial ex- periments in cultured SMCs to dissect the signaling leading to necrotic cell death (Aim 1B); and evaluate SMC mitochondrial damage and necrotic cell death in patient tissues by electron microscopy analysis (Aim 1B). Aim 2. We will test the hypothesis that cGAS and RIP3-mediated SMC necroptosis are critically involved in aortic SMC injury, aortic destruction, inflammation, biomechanical failure, and ATAAD development by comparing these aspects in WT mice, cGas-/- mice, Rip3-/- mice, and SMC specific Rip3 knockout mice in a sporadic ATAAD model induced by HFD/Ang II challenge. Aim 3. We will test the hypothesis that pharmacologically re- ducing mitochondrial damage or necroptosis will prevent aortic destruction and disease progression in our spo- radic ATAAD mouse model. The proposed research is significant because it will not only provide novel insight of the molecular mechanisms of sporadic ATAAD development, but also test new treatments for preventing sporadic ATAAD formation and its fatal sequelae. The novel mechanisms of mitochondrial damage-induced SMC necroptosis will also have broad implications in many other cardiovascular diseases.